Abstract
The aim of the present study was to explore the role of miR‑493-5p in liver cancer tissues and cell lines, and its effect on cell behavioral characteristics. The expression of miR-493-5p was detected by reverse transcription-quantitative polymerase chain reaction(RT-qPCR) in liver cancer tissues and cell lines (hepatic cell line HL-7702 and the liver cancer cell lines HCCC-9810, HuH-7 and HepG2). In addition, the mechanism by which miR-493-5p mediates its effects was analyzed via the transfection of miR-493-5p mimic and negative control miRNA into HepG2cells. The viability, proliferation, apoptosis and invasion of the cells were analyzed using MTT assay, flow cytometry and Transwell chamber experiments. Furthermore, the effect of miR-493-5p on the expression of vesicle associated membrane protein2(VAMP2) was assayed using a dual-luciferase reporter system, and VAMP2 protein levels were determined by western blot analysis. In addition, following the cotransfection of HepG2 cells with pcDNA3.1‑VAMP2 plasmid and miR‑493-5p mimic, the role of miR-493-5p as a regulator of VAMP2 was evaluated using MTT assay, flow cytometry and Transwell chamber experiments. RT-qPCR analysis indicated that the expression of miR-493-5p in liver cancer tissues and cell lines was decreased significantly compared with that in adjacent normal liver tissues and normal liver cell lines, respectively. Compared with the control group, the cells transfected with miR-493-5p mimic (the miR-493-5p overexpression group) exhibited reduced cell viability, a reduced percentage of cells in the Sphase and an increased percentage of apoptotic cells. In addition, fewer cells passed through the Transwell membrane in the miR-493-5p overexpression group compared with the control group. In the dual-luciferase reporter assay, luciferase activity in the miR‑493-5p overexpression group was attenuated compared with that in the control group. Inaddition, western blot analysis indicated that the VAMP2 protein levels in the miR‑493-5p overexpression group were lower than those in the control group. Furthermore, in cells overexpressing miR-493-5p and VAMP2 simultaneously, the biological behavior of the cells, including cell viability, cell cycle and cell invasiveness, was significantly rescued compared with that of the control group transfected with miR‑493-5p alone. In conclusion, miR-493-5p is indicated to be a tumor suppressor gene, and is downregulated in human liver cancer. miR-493-5p overexpression promotes cell apoptosis and inhibits the proliferation and migration of liver cancer cells by negatively regulating the expression of VAMP. These observations suggest the potential of treating liver cancer by the overexpression of microRNA-493-5p.
Highlights
Liver cancer ranks sixth for incidence and third for mortality among the most common cancers worldwide [1]
The results of reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis indicated that the expression of miR-493-5p in the HCC tissue samples was significantly lower than that in the adjacent normal tissues (P
The expression of miR-493-5p in the liver cancer cell lines HCCC‐9810, HuH-7 and HepG2 was reduced significantly compared with that in the HL-7702 hepatic cells (P
Summary
Liver cancer ranks sixth for incidence and third for mortality among the most common cancers worldwide [1]. In 2014, it was reported that among 87,988 malignant liver and intrahepatic cancer diagnoses reported during 2000-2010 in SEER 18 registries, 63,735 (72%) were classified as HCCs in USA. Between 2006 and 2010, US liver cancer mortality rates increased with age in all racial/ethnic groups [2]. In the USA, the incidence of liver cancer has been rising faster than that of other malignant cancers [4]. The occurrence of liver cancer is closely correlated with infection by hepatitis B virus (HBV), and 50% of cases result from HBV infection, the infection rate of which is high [5,6]. Great advances in the surgical therapy and other treatments for liver cancer have been achieved, the 5-year survival rate is only 5-6% and the poor prognosis is a cause of severe economic loss and a heavy burden for society [7,8,9]
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